Regulation and Function of Myocardin in Vascular Pathobiology

描述（由申请方提供）：血清反应因子（SRF）和肌心素（MYOCD）构成了一种分子触发开关，用于激活一组含有SRF结合CArG盒的血管平滑肌细胞（VSMC）细胞收缩和离子通道基因。我们首先发现MYOCD水平（而不是SRF）与VSMC分化程度相关。由于VSMC的分化程序是不固定的，并在许多疾病过程中的MYOCD水平变化的适应，我们一直有兴趣了解MYOCD表达的调节和它的功能作为一个稳态开关的保存一个静止的，收缩状态的VSMC。因此，我们在一系列论文中报道了MYOCD（a）足以通过收缩和离子通道基因表达的SRF依赖性变化赋予VSMC收缩能力;（B）完全抑制骨骼肌分化程序;（c）控制越来越多的microRNA，包括microRNA 143/145基因;（d）由TGF β 1阳性诱导。1通过p38 MAPK依赖性途径。初步数据进一步证明了转录抑制因子和新microRNA的顺式元件，以及首次显示MYOCD表达对血管闭塞性疾病、脂质摄取和炎症标志物表达的有利影响的功能数据。总的来说，我们越来越多的工作作为一个关键的基础，以测试的假设，MYOCD是一个正常的VSMC分化的稳态开关。本论文将通过一系列相互关联的特异性目的设计来阐明MYOCD表达的转录和转录后调控以及MYOCD在损伤诱导的新生内膜扩张和动脉粥样硬化形成中的血管重塑中的作用。目的1：利用细菌人工染色体转基因小鼠研究MYOCD的顺式作用元件的功能。在目标2中，新型MYOCD功能丧失和获得小鼠将直接评估这种强大的辅因子在实验性血管疾病过程中的作用，包括对VSMC炎症，增殖和转分化状态的影响。在目标3中，将进行基因组研究，其整合了我们已经产生的人和小鼠CArGome数据（> 84，000个CArG盒）与VSMC的RNA-seq，其中MYOCD在SRF存在或不存在的情况下表达。这样的分析将揭示CAr基因组的一个子集，其响应于MYOCD以及血管疾病中的新的SRF非依赖性MYOCD靶基因。因此，计划中的研究将产生新的见解，在正常的出生后发育和血管疾病过程中的体内调控MYOCD表达，以及新的信息相关的MYOCD作为一个可能的抑制剂与物理损伤和动脉粥样硬化性疾病相关的血管重塑。这些研究的结果将具有巨大的应用，用于设计新的治疗策略以对抗急性和慢性血管疾病以及可能改变MYOCD表达/活性的其他疾病（例如，哮喘、高血压、阿尔茨海默病）。

DESCRIPTION (provided by applicant): Serum response factor (SRF) and Myocardin (MYOCD) constitute a molecular trigger switch for the activation of a battery of vascular smooth muscle cell (VSMC) cyto-contractile and ion channel genes containing SRF- binding CArG boxes. We first showed that levels of MYOCD (but not SRF) correlate with the degree of VSMC differentiation. Since the VSMC differentiation program is not fixed and subject to adaptation in a number of disease processes where levels of MYOCD change, we have been interested in understanding the regulation of MYOCD expression and its function as a homeostatic switch for the preservation of a quiescent, contractile state in VSMC. Accordingly, we have reported in a series of papers that MYOCD (a) is sufficient for conferring VSMC contractile competence through SRF-dependent changes in contractile and ion channel gene expression; (b) completely represses the program of skeletal muscle differentiation; (c) controls a growing number of microRNAs, including the microRNA143/145 gene; and (d) is positively induced by TGF¿1 through a p38MAPK-dependent pathway. Preliminary data further demonstrate cis elements for a transcriptional repressor and new microRNA as well as functional data showing, for the first time, the favorable effects of MYOCD expression on vascular occlusive disease, lipid uptake, and inflammatory marker expression. Collectively, our growing body of work serves as a critical foundation to test the hypothesis that MYOCD is a homeostatic switch for normal VSMC differentiation. This thesis will be tested through a series of inter-related specific aims designe to elucidate transcriptional and post-transcriptional control of MYOCD expression and the role of MYOCD in vascular remodeling during injury-induced neointimal expansion and atherogenesis. In Aim 1, bacterial artificial chromosome transgenic mice will elucidate the function of novel cis-acting elements leading to activation or repression of MYOCD. In Aim 2, novel MYOCD loss- and gain-of-function mice will directly assess the role of this powerful cofactor in experimental vascular disease processes, including effects on VSMC inflammatory, proliferative, and transdifferentiative states. In Aim 3, genomic studies integrating human and mouse CArGome data we have generated (> 84,000 CArG boxes) with RNA-seq of VSMC where MYOCD is expressed in the absence or presence of SRF will be carried out. Such an analysis will reveal a subset of the CArGome that is responsive to MYOCD as well as new SRF-independent MYOCD target genes of import in vascular disease. Thus, the planned studies will yield new insight into the in vivo regulation of MYOCD expression during normal postnatal development and in vascular disease processes as well as novel information related to MYOCD as a likely inhibitor of vascular remodeling associated with physical injury and atherosclerotic disease. The results of these studies will have enormous applications for devising new therapeutic strategies to combat acute and chronic vascular diseases and perhaps other diseases where MYOCD expression/activity may be altered (e.g., asthma, hypertension, Alzheimer's disease).